Abscisic acid sensor RCAR7/PYL13, specific regulator of protein phosphatase coreceptors

Abstract

The plant hormone abscisic acid (ABA) acts both as a developmental signal and as an integrator of environmental cues such as drought and cold. ABA perception recruits an ABA-binding regulatory component [regulatory component of ABA receptor (RCAR)/ PYR1/PYL] and an associated protein phosphatase 2C (PP2C). Phytohormone binding inactivates the phosphatase activity of the coreceptor, permitting phosphorelay of the ABA signal via downstream protein kinases. RCARs and PP2C coreceptors are represented by small protein families comprising 14 and 9 members in Arabidopsis, respectively. The specificity of the RCAR-PP2C interaction and the constraints contributing to specific combinations are poorly understood. In this contribution, we analyzed RCAR7/PYL13, which is characterized by three variant amino acid residues in the conserved ABA-binding pocket. RCAR7 regulated the phosphatase activity of the PP2Cs ABI1, ABI2, and PP2CA in vitro at nanomolar ABA levels; however, it was unable to regulate the structurally related hypersensitive to ABA 1 (HAB1). Site-directed mutagenesis of HAB1 established ABA-dependent regulation by RCAR7. Conversion of the noncanonical amino acid residues of RCAR7 into the consensus ABA-binding pocket did not perceptibly change receptor function. Ectopic expression of RCAR7 in Arabidopsis resulted in ABA hypersensitivity affecting gene regulation, seed germination, and stomatal closure. The RCAR7 loss-of-function mutant revealed no changes in ABA responses, similar to the RCAR9 knockout line, whereas the combined deficiency of RCAR7 and RCAR9 resulted in ABA-insensitive seed germination. The study shows a role of RCAR7 in early plant development, proves its ABA receptor function, and identifies structural constraints of RCAR7-PP2C interaction.

Combinatorial interaction network of abscisic acid receptors and coreceptors from Arabidopsis thaliana

“…In the same context, selective discrimination of PP2Cs by specific RCARs such as RCAR7/PYL13 point to RCAR-specific features. In fact, analysis-of-specificity determinants for RCARs (30,33,45,46) and PP2C HAB1 (32) and AHG1 (this study) showed that both receptor components have evolved key-lock features. RCAR7 did not rescue ABI1-, AHG1-, and HAB1-inhibited ABA signaling in this study in agreement with previous analyses (32,35).…”

“…In fact, analysis-of-specificity determinants for RCARs (30,33,45,46) and PP2C HAB1 (32) and AHG1 (this study) showed that both receptor components have evolved key-lock features. RCAR7 did not rescue ABI1-, AHG1-, and HAB1-inhibited ABA signaling in this study in agreement with previous analyses (32,35). Characterization of the RCAR7-specific repulsion by HAB1 revealed that changes in two amino acid residues of HAB1 located at the RCAR interaction surface confer RCAR7 binding and regulation (32).…”

“…In the presence of ABA, ABA signaling is stimulated by ectopic expression of RCARs; however, the response was found to be quite variable (13,27,32,34,35). The variations might be caused by different ABA affinities and ABA-independent action of RCARs (28) or by the abundance of target PP2Cs and RCARs.…”

“…2 A-C). The regulation of the ABA signal pathway by a PP2C-RCAR pair can be evaluated by comparing the ABA response of cells inhibited by ectopic expression of a single PP2C with cells in which a specific RCAR was coexpressed in the presence and absence of exogenous ABA (32). To exemplify the potential of the assay system, ABA signaling was reduced to ∼10% by ectopic expression of HAB1 and activated by coexpression of RCAR12 (Fig.…”

Depletion of abscisic acid levels in roots of flooded Carrizo citrange (Poncirus trifoliata L. Raf. × Citrus sinensis L. Osb.) plants is a stress-specific response associated to the differential expression of PYR/PYL/RCAR receptors

“…This implies a fine adjustment of the magnitude of the physiological response and requires a more complex regulatory system than the sole presence/absence of ABA. Indeed, there are 14 loci encoding for ABA receptors in Arabidopsis, each of them having different intrinsic affinities for the hormone and could also bind to specific PP2Cs (Dupeux et al 2011;Santiago et al 2012;Antoni et al 2013;Fuchs et al 2014). Besides the fine-tuning of the ABA signal, modulation of the endogenous hormone levels is associated to the specific environmental situation and the stress pressure, which adds further complexity to the model (Seiler et al 2011;Romero et al 2012b; Baron et al 2012;De Ollas et al 2013).…”